The long-term objectives of this research program are to help define the genetic, structural and pathogenic basis of meningococcal endotoxin (lipoligosaccharide[LOS]). The recent dramatic increases in meningococcal infections in part of the United States and the emergency of penicillin-resistant meningococcal, emphasize pathogenesis. The principle investigator s approach utilized novel genetic techniques for meningococcal(e.g., Tn916 mutagenesis), biochemical and physical analysis of LOS structure; and human tissue assays to assemble a detailed model of meningococcal LOS and to provide insights into how Los influences pathogenesis. This application is directed at understanding the structural-functional relationship of the meningococcal LOS inner core, Flu (GlcNAc) Hep2KD02LipidA. LOS inner core structure is predicted to influence LOS assembly, human immune recognition of the meningococcus and interactions with soluble and cellular human proteins.
In Specific Aim 1, genetic determinants that are responsible for LOS inner core oligosaccharide chain synthesis and heterogeneity in serogroup B meningococcal will be characterized: (1) by investigation of the ace (alpha chain elongation) operon which contains the UDP-Glu: heptosyl (ll) lipooligosaccharide alpha 1, 2 N- acetylglucosamine transferase (rfaK) and the putative UDP-Glu heptosyl (1) lipooligosaccharide beta 1, 4 glucosyltransferase (IgtF); (2) by study of alpha chain initiation, and additions of phosphate derivatives and saccharides to Hepll; and (3) by understanding the deep rough Tn916 mutant 469 (KD02LipidA) at both genetic and structural levels.
In Specific Aim 2, the role of Los in two important events that occur during group B meningococcal infection will be investigated. They propose that the LOS inner core, in particular the Hepll structure, facilitates group B meningococcal entry into human nasopharyngeal epithelial cells. In addition, genetically and structurally defined LOS mutants will be used to determine how expression of hep2KD02-Lipid A LOS enhances sensitivity of encapsulated meningococcal to killing by classical pathway complement-mediate mechanisms of human serum. Such data has application to the design of meningococcal vaccines currently in development (e.g. inclusion of truncated LOS molecules in outer membrane vesicle vaccines), and should provide insights into the unique features of meningococcal endotoxin.
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